Coupling: No changes needed from standard method recommended by synthesizer manufacturer. Monomers that allow for UltraMILD deprotection are recommended (Catalog Numbers: dA: 10-1601-xx, dC: 10-1015-xx, dG: 10-1621-xx, dT: 10-1030-xx.) To avoid any exchange of the iPr-Pac group on the dG with acetyl, use the UltraMild Cap Mix A (40-4210-xx/ 40-4212-xx).
If UltraMild Monomers were used: 0.05M Potassium Carbonate in Methanol, 4hrs at Room Temp.
DMT-Off Synthesis: Add 6uL acetic acid per mL 0.05M potassium carbonate in methanol. Dry under vacuum. Desalt per standard procedures.
DMT-On Synthesis: Use a Glen-Pak Purification Cartridge (60-5100-XX/60-5200-XX) to purify and desalt the oligonucleotide. Dilute the 0.05M potassium carbonate in methanol with 100mg/mL Sodium Chloride to a final concentration of 1:4(v/v). Load onto the prepped Glen-Pak Cartridge and purify as normal.
If Standard Monomers were used:
1) Prepare a solution of 10% DBU in anhydrous methanol. DBU (1,8-Diazabicyclo[5.4.0]undec-7-ene) is available from Aldrich (139009-25G) as is anhydrous methanol (322415-100ML).
2) Transfer the CPG to a clean, dry glass vial and add 1mL of the 10% DBU solution.
3) Seal the vial and leave for 5 days in the dark at room temperature.
4) Transfer the contents to a micro centrifuge tube and reduce under vacuum to a small volume.
5) Add 1mL of 10mM aqueous sodium hydroxide solution and desalt or purify the oligonucleotide using standard procedures.
Storage: Refrigerated storage, maximum of 2-8°C, dry
The table below shows pack size data and, for solutions, dilutions and approximate couplings based on normal priming procedures. Please link for more detailed usage information with the various synthesizers.